Intra-Vκ Cluster Recombination Shapes the Ig Kappa Locus Repertoire

During V(D)J recombination RAG proteins introduce DNA double strand breaks (DSBs) adjacent to conserved recombination signal sequences (RSS) that contain either 12- or 23-nucleotide spacer regions. Coordinated cleavage following the “12/23 rule” predicts that DSBs at variable (V) gene segments should not exceed the level of breakage at joining (J) segments, thereby ensuring that V regions do not engage in undesirable recombination events with one another. Here we report abundant RAG dependent DSBs at a multitude of Vκ gene segments within the Igk locus independent of V-J rearrangement. We discover that a large fraction of Vκ gene segments are flanked not only by a bone-fide 12 spacer, but also an overlapping, 23 spacer flipped RSS. These compatible pairs of RSS mediate recombination and deletion inside the Vκ cluster even in the complete absence of Jκ gene segments, and support a novel recombination center (RC) independent of the conventional Jκ-centered RC. We propose a model that explains Vκ gene segment usage by taking into account not only the probability of V-to-J rearrangement but also the surprisingly frequent, evolutionarily conserved intra-Vκ cluster recombination events. These findings shed light on the diverse molecular strategies that shape the primary antigen receptor repertoires.